Fatigue, excessive sleepiness, excess sleep and sleep disturbances are presenting symptoms in nearly all infectious diseases. The broad objective of this grant proposal is to characterize the molecular mechanisms responsible for changes in sleep induced by influenza virus. These investigators hypothesize that viral double-stranded (ds)RNA induces an upregulation of physiological somnogenic cytokines and de novo synthesis of other cytokines that amplify physiological sleep mechanisms, which include activation of nitric oxide synthase (NOS). Substantial preliminary data support this broad hypothesis. The model used for these sleep studies is A/PR/8/34-H1N1 influenza virus infection in the mouse. PR8 causes a pneumonitis accompanied by early onset of sleep responses. In the first Specific Aim, cytokine profiles induced by influenza virus or by the synthetic dsRNA,poly I:C in lungs will be compared. It is expected that the appearance of viral dsRNA will precede an upregulation of cytokine mRNA and that poly I:C will induce a similar pattern of cytokine expression. Further, the effects of antibody neutralization of viral dsRNA and the role of cytokine priming will be determined. Specific Aim #2 focuses on cytokine production in the brain and its involvement in viral-induced sleep responses. Three knockout strains of mice, each lacking receptors of one or more cytokines, will be infected and sleep responses determined. The ability of these mice to cope with the infectious challenge is expected to be compromised and their sleep responses blunted. Control mice similarly infected will be examined for cytokine mRNA expression in various area of brain. Brain expression of cytokines is expected to precede sleep responses and be localized to the hypothalamus and hippocampus. In Specific Aim #3 the role that NOS plays in viral-induced sleep responses will be determined. Three different NOS-knockout strains of mice (lacking NOS-1, -2 and -3, respectively) will be inoculated with influenza virus and sleep responses will be determined. The localization of the expression of NOS mRNAs in brain of similarly infected control mice will also be determined. It is anticipated that NOS-2 knockouts will be the more severely impaired mice and that viral-induced sleep responses will be attenuated in both NOS-1 and NOS-2 knockout mice. Viral-induced changes in NOS expression are expected to be localized in brain and be evident in lung tissue. The anticipated results will greatly aid our understanding of the molecular mechanisms involved in viral-induced sleep responses as well as physiological sleep mechanisms.